Both insulin-dependent (Type I) and non-insulin-dependent (Type II) diabetes are characterized by alterations in insulin action and glucose metabolism in peripheral tissues including muscle, fat and liver. Many of these alterations are secondary to the altered metabolism which accompanies both types of diabetes. In addition, in Type II diabetes there is considerable evidence for some primary alteration(s) in insulin action. These primary and secondary alterations in insulin action are mediated via changes in both expression of genes at the mRNA level and by post-translational modification of proteins. The long-term goal of this project is to identify changes in gene expression at the level of muscle in humans with diabetes mellitus using techniques which are not limited to studying previously identified genes, to determine which of these are secondary to the metabolic abnormalities of diabetes (i.e., "diabetes-related" changes in gene expression which can be present in both Type I and Type II diabetes) versus those which might be primary defects (i.e., unique "diabetogenes") related to the insulin resistance of Type II diabetes. In preliminary experiments, we have shown that this can be achieved using subtraction libraries produced from skeletal muscle cDNA screened with subtractive probes. Thus far, 11 diabetes-related genes have been identified including three not previously in GenBank, one of which is uniquely altered in muscle of Type II diabetics and is a new member of the ras-GTPase family. In the proposed experiments we will: 1) Identify additional alterations in gene expression in human muscle from individuals with Type I and Type II diabetes by subtractive cloning and screening with differential and subtractive probes; 2) Determine which of these alterations are common to both types of diabetes, and hence probably secondary to the metabolic abnormalities in diabetes, versus those that are limited to Type II diabetes which may be primary genetic defects in this disease or reflect specific alterations due to insulin resistance; 3) Characterize the candidate diabetes-related genes with respect to tissue distribution for expression, size and number of mRNA species, relationship to metabolic control and complications; and 4) Determine the exact nature of the diabetes-related genes by sequencing of the cDNA clones, comparison to known sequences in gene data banks, and ultimately by expression of the protein encoded by the gene and/or development of antibodies to the protein.